Berberine inhibits intracellular Ca2+ signals in mouse pancreatic acinar cells through M3 muscarinic receptors: Novel target, mechanism, and implication.

Berberine, a natural isoquinoline alkaloid, exhibits a variety of pharmacological effects, but the pharmacological targets and mechanisms remain elusive. Here, we report a novel finding that berberine inhibits acetylcholine (ACh)-induced intracellular Ca2+ oscillations, mediated through an inhibition of the muscarinic receptor subtype 3 (M3) receptor. Patch-clamp recordings and confocal Ca2+ imaging were applied to acute dissociated pancreatic acinar cells prepared from CD1 mice to examine the effects of berberine on ACh-induced Ca2+ oscillations. Whole-cell patch-clamp recordings showed that berberine (from 0.1 to 10 µM) reduced ACh-induced Ca2+ oscillations in a concentration-dependent manner, and this inhibition also depended on ACh concentrations. The inhibitory effect of berberine neither occurred in intracellular targets nor extracellular cholecystokinin (CCK) receptors, chloride (Cl-) channels, and store-operated Ca2+ channels. Together, the results demonstrate that berberine directly inhibits the muscarinic M3 receptors, further confirmed by evidence of the interaction between berberine and M3 receptors in pancreatic acinar cells.

Electrophysiological Phenotypes of Hippocampal Synaptic and Network Functions in Cannabinoid Receptor 2 Knockout Mice.

Background: The cannabinoid receptor 2 (CB2R), a cannabinoid receptor primarily expressed in immune cells, has been found in the brain, particularly in the hippocampus, where it plays crucial roles in modulating various neural functions, including synaptic plasticity, neuroprotection, neurogenesis, anxiety and stress responses, and neuroinflammation. Despite this growing understanding, the intricate electrophysiological characteristics of hippocampal neurons in CB2R knockout (CB2R KO) mice remain elusive. Aim and Methods: This study aimed to comprehensively assess the electrophysiological traits of hippocampal synaptic and network functions in CB2R KO mice. The focus was on aspects such as synaptic transmission, short- and long-term synaptic plasticity, and neural network synchrony (theta oscillations). Results: Our findings unveiled multiple functional traits in these CB2R KO mice, notably elevated synaptic transmission in hippocampal CA1 neurons, decreased both synaptic short-term plasticity (paired-pulse facilitation) and long-term potentiation (LTP), and impaired neural network synchronization. Conclusion: In essence, this study yields insightful revelations about the influence of CB2Rs on hippocampal neural functions. By illuminating the electrophysiological modifications in CB2R KO mice, our research enriches the comprehension of CB2R involvement in hippocampal function. Such insights could hold implications for advancing our understanding of the neural mechanisms under the influence of CB2Rs within the brain.

Endogenous Amyloid-formed Ca2+-permeable Channels in Aged 3xTg AD Mice.

Alzheimer's disease (AD), the leading cause of dementia, is characterized by the accumulation of beta-amyloid peptides (Aß). However, whether Aß itself is a key toxic agent in AD pathogenesis and the precise mechanism of Aß-elicited neurotoxicity are still debated. Emerging evidence demonstrates that the Aß channel/pore hypothesis could explain Aß toxicity, because Aß oligomers are able to disrupt membranes and cause edge-conductivity pores that may disrupt cell Ca2+ homeostasis and drive neurotoxicity in AD. However, all available data to support this hypothesis have been collected from "in vitro" experiments using high concentrations of exogenous Aß. It is still unknown whether Aß channels can be formed by endogenous Aß in AD animal models. Here, we report an unexpected finding of the spontaneous Ca2+ oscillations in aged 3xTg AD mice but not in age-matched wild-type mice. These spontaneous Ca2+ oscillations are sensitive to extracellular Ca2+, ZnCl2, and the Aß channel blocker Anle138b, suggesting that these spontaneous Ca2+ oscillations in aged 3xTg AD mice are mediated by endogenous Aß-formed channels.

Impact of the Cannabinoid System in Alzheimer's Disease.

Cannabinoids are compounds isolated from cannabis and are also widely present in both nervous and immune systems of animals. In recent years, with in-depth research on cannabinoids, their clinical medicinal value has been evaluated, and many exciting achievements have been continuously accumulating, especially in the field of neurodegenerative disease. Alzheimer's disease is the most common type of neurodegenerative disease that causes dementia and has become a global health problem that seriously impacts human health today. In this review, we discuss the therapeutic potential of cannabinoids for the treatment of Alzheimer's disease. How cannabinoids act on different endocannabinoid receptor subtypes to regulate Alzheimer's disease and the roles of the endocannabinoid system in Alzheimer's disease are outlined, and the underlying mechanisms are discussed. Finally, we summarize the most relevant opportunities of cannabinoid pharmacology related to Alzheimer's disease and discuss the potential usefulness of cannabinoids in the clinical treatment of Alzheimer's disease.

Genome-Wide Identification, Characterization, and Expression Analysis of Tubby-like Protein (TLP) Gene Family Members in Woodland Strawberry (Fragaria vesca).

Tubby-like proteins (TLPs) play important roles in plant growth and development and in responses to abiotic stress. However, TLPs in strawberry remain poorly studied. In this study, eight TLPs were identified in woodland strawberry (Fragaria vesca subspecies vesca 'Ruegen'). Protein structure analysis revealed that the structure of FvTLPs is highly conserved, but evolutionary and gene structure analyses revealed that the evolutionary pattern of FvTLP family members differs from that of their orthologous genes in Arabidopsis, poplar, and apple. Subcellular localization assays revealed that FvTLPs were localized to the nucleus and plasma membrane. FvTLPs showed no transcriptional activity. Yeast two-hybrid assays revealed that FvTLPs interact with specific FvSKP1s. The expression patterns of FvTLPs in different tissues and under various abiotic stresses (salt, drought, cold, and heat) and hormone treatments (ABA (abscisic acid) and MeJA (methyl jasmonate)) were determined. The expression patterns of FvTLPs indicated that they play a role in regulating growth and development and responses to abiotic stress in F. vesca. The GUS (beta-glucuronidase) activity of FvTLP1pro::GUS plants in GUS activity assays increased under salt and drought stress and abscisic acid treatment. The results of this study provide new insights into the molecular mechanisms underlying the functions of TLPs.

FaAKR23 Modulates Ascorbic Acid and Anthocyanin Accumulation in Strawberry (Fragaria × ananassa) Fruits.

Strawberry (Fragaria × ananassa) fruits are rich in ascorbic acid (AsA) and anthocyanin, which are essential antioxidants for human health. However, the underlying regulatory mechanism of these antioxidant accumulation, especially AsA accumulation in strawberry fruits, remains largely unknown. In this study, we identified FaAKR23 was a regulator of AsA and anthocyanin accumulation. We transiently expressed FaAKR23 in strawberry fruits and conducted metabolic and molecular analyses to explore the role of FaAKR23 in AsA and anthocyanin accumulation. Transient silencing of FaAKR23 (FaAKR23-RNAi) in strawberry fruits significantly decreased the AsA and anthocyanin contents compared with control (empty vector-RNAi, EV-RNAi). Correspondingly, expression of some structural genes and regulatory factors involved in these two antioxidants' accumulation was dramatically repressed. In addition, transcriptome analysis of EV-RNAi and FaAKR23-RNAi fruits suggested that FaAKR23 was also involved in starch and sucrose metabolism as well as plant-pathogen interaction. Overall, these results not only provide the coordinated regulatory function of FaAKR23 on AsA and anthocyanin accumulation but also offer a promising candidate gene for strawberry breeding with high antioxidants.

Expression analysis and functional characterization of tomato Tubby-like protein family.

Tubby-like protein (TLP) plays an important role in plant growth and development. In this investigation, the characteristics of 11 members in the SlTLP family were studied. SlTLP genes were classified into two subgroups, and the members containing the F-box domain were renamed SlTLFPs. Subcellular localization indicated that most of the SlTLPs were localized in the nucleus. Expression pattern analysis revealed that eight genes (SlTLFP1, 3, 5, 7-10, and SlTLP11) showed differential expression across various tissues, while SlTLFP2, 4, and 6 were widely expressed in all the organs tested. Most SlTLP genes were induced by biotic and abiotic stress treatments such as Botrytis cinerea, temperature, MeJA, and ABA. TLP proteins in tomato have no transcriptional activation activity, and most members with an F-box domain could interact with SUPPRESSOR OF KINETOCHORE PROTEIN 1 (SlSkp1) or Cullin1 (Cul1) or both. Experiments on CRISPR edited SlTLFP8 showed that the N-terminal F-box domain was necessary for its function such as DNA ploidy and stomata size regulation. Our findings suggested that the F-box domain interacts with Skp1 and Cul1 to form the SCF complex, suggesting that SlTLFPs, at least SlTLFP8, function mainly through the F-box domain as an E3 ligase.

Combined transcriptomic and metabolomic analysis reveals a role for adenosine triphosphate-binding cassette transporters and cell wall remodeling in response to salt stress in strawberry.

Strawberry (Fragaria × ananassa Duch) are sensitive to salt stress, and breeding salt-tolerant strawberry cultivars is the primary method to develop resistance to increased soil salinization. However, the underlying molecular mechanisms mediating the response of strawberry to salinity stress remain largely unknown. This study evaluated the salinity tolerance of 24 strawberry varieties, and transcriptomic and metabolomic analysis were performed of 'Sweet Charlie' (salt-tolerant) and 'Benihoppe' (salt-sensitive) to explore salt tolerance mechanisms in strawberry. Compared with the control, we identified 3412 differentially expressed genes (DEGs) and 209 differentially accumulated metabolites (DAMs) in 'Benihoppe,' and 5102 DEGs and 230 DAMs in 'Sweet Charlie.' DEGs Gene Ontology (GO) enrichment analyses indicated that the DEGs in 'Benihoppe' were enriched for ion homeostasis related terms, while in 'Sweet Charlie,' terms related to cell wall remodeling were over-represented. DEGs related to ion homeostasis and cell wall remodeling exhibited differential expression patterns in 'Benihoppe' and 'Sweet Charlie.' In 'Benihoppe,' 21 ion homeostasis-related DEGs and 32 cell wall remodeling-related DEGs were upregulated, while 23 ion homeostasis-related DEGs and 138 cell wall remodeling-related DEGs were downregulated. In 'Sweet Charlie,' 72 ion homeostasis-related DEGs and 275 cell wall remodeling-related DEGs were upregulated, while 11 ion homeostasis-related DEGs and 20 cell wall remodeling-related DEGs were downregulated. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses showed only four KEGG enriched pathways were shared between 'Benihoppe' and 'Sweet Charlie,' including flavonoid biosynthesis, phenylalanine metabolism, phenylpropanoid biosynthesis and ubiquinone, and other terpenoid-quinone biosynthesis. Integrating the results of transcriptomic and metabolomics analyses showed that adenosine triphosphate-binding cassette (ABC) transporters and flavonoid pathway genes might play important roles in the salt stress response in strawberry, and DAMs and DEGs related to ABC transporter and flavonoid pathways were differentially expressed or accumulated. The results of this study reveal that cell wall remodeling and ABC transporters contribute to the response to salt stress in strawberry, and that related genes showed differential expression patterns in varieties with different salt tolerances. These findings provide new insights into the underlying molecular mechanism of strawberry response to salt stress and suggest potential targets for the breeding of salt-tolerant strawberry varieties.

Chromosome-level genome assembly of Fragaria pentaphylla using PacBio and Hi-C technologies.

Fragaria pentaphylla, a wild diploid quinquefoliolate species of Fragaria, is native to Southwest China. It has two morphs of red and white fruit color in nature and has characteristics of unique fragrance and resistance, which made it not only a valuable breeding material but also a potential model plant for molecular function researches. Here, we generate a high-quality chromosome-level genome assembly of a F. pentaphylla accession, BAAFS-FP039 employing a combination of PacBio Long-Read Sequencing, Illumina Short-Read Sequencing, and Hi-C Sequencing. The assembled genome contained 256.74 Mb and a contig N50 length of 32.38 Mb, accounting for 99.9% of the estimated genome (256.77 Mb). Based on Hi-C data, seven pseudo-chromosomes of F. pentaphylla-FP039 genome were assembled, covering 99.39% of the genome assembly. The genome was composed of 44.61% repetitive sequences and 29,623 protein-coding genes, 97.62% of protein-coding genes could be functionally annotated. Phylogenetic and chromosome syntenic analysis revealed that F. pentaphylla-FP039 was closely related to F. nubicola. This high-quality genome could provides fundamental molecular resources for evolutionary studies, breeding efforts, and exploring the unique biological characteristics of F. pentaphylla.

Genome-Wide Analysis of Ascorbic Acid Metabolism Related Genes in Fragaria × ananassa and Its Expression Pattern Analysis in Strawberry Fruits.

Ascorbic acid (AsA) is an important antioxidant for scavenging reactive oxygen species and it is essential for human health. Strawberry (Fragaria × ananassa) fruits are rich in AsA. In recent years, strawberry has been regarded as a model for non-climacteric fruit ripening. However, in contrast to climacteric fruits, such as tomato, the regulatory mechanism of AsA accumulation in strawberry fruits remains largely unknown. In this study, we first identified 125 AsA metabolism-related genes from the cultivated strawberry "Camarosa" genome. The expression pattern analysis using an available RNA-seq data showed that the AsA biosynthetic-related genes in the D-mannose/L-galactose pathway were downregulated remarkably during fruit ripening which was opposite to the increasing AsA content in fruits. The D-galacturonate reductase gene (GalUR) in the D-Galacturonic acid pathway was extremely upregulated in strawberry receptacles during fruit ripening. The FaGalUR gene above belongs to the aldo-keto reductases (AKR) superfamily and has been proposed to participate in AsA biosynthesis in strawberry fruits. To explore whether there are other genes in the AKR superfamily involved in regulating AsA accumulation during strawberry fruit ripening, we further implemented a genome-wide analysis of the AKR superfamily using the octoploid strawberry genome. A total of 80 FaAKR genes were identified from the genome and divided into 20 subgroups based on phylogenetic analysis. These FaAKR genes were unevenly distributed on 23 chromosomes. Among them, nine genes showed increased expression in receptacles as the fruit ripened, and notably, FaAKR23 was the most dramatically upregulated FaAKR gene in receptacles. Compared with fruits at green stage, its expression level increased by 142-fold at red stage. The qRT-PCR results supported that the expression of FaAKR23 was increased significantly during fruit ripening. In particular, the FaAKR23 was the only FaAKR gene that was significantly upregulated by abscisic acid (ABA) and suppressed by nordihydroguaiaretic acid (NDGA, an ABA biosynthesis blocker), indicating FaAKR23 might play important roles in ABA-mediated strawberry fruit ripening. In a word, our study provides useful information on the AsA metabolism during strawberry fruit ripening and will help understand the mechanism of AsA accumulation in strawberry fruits.

A tubby-like protein CsTLP8 acts in the ABA signaling pathway and negatively regulates osmotic stresses tolerance during seed germination.

BACKGROUND: TLPs (Tubby-like proteins) are widespread in eukaryotes and highly conserved in plants and animals. TLP is involved in many biological processes, such as growth, development, biotic and abiotic stress responses, while the underlying molecular mechanism remains largely unknown. In this paper we characterized the biological function of cucumber (Cucumis sativus L.) Tubby-like protein 8 (CsTLP8) in Arabidopsis. RESULTS: In cucumber, the expression of the tubby-like protein CsTLP8 was induced by NaCl treatment, but reduced by PEG (Polyethylene Glycol) and ABA (Abscisic Acid) treatment. Subcellular localization and transcriptional activation activity analysis revealed that CsTLP8 possessed two characteristics of classical transcription factors: nuclear localization and trans-activation activity. Yeast two-hybrid assay revealed interactions of CsTLP8 with CsSKP1a and CsSKP1c, suggesting that CsTLP8 might function as a subunit of E3 ubiquitin ligase. The growth activity of yeast with ectopically expressed CsTLP8 was lower than the control under NaCl and mannitol treatments. Under osmotic and salt stresses, overexpression of CsTLP8 inhibited seed germination and the growth of Arabidopsis seedlings, increased the content of MDA (Malondialdehyde), and decreased the activities of SOD (Superoxide Dismutase), POD (Peroxidase) and CAT (Catalase) in Arabidopsis seedlings. Overexpression of CsTLP8 also increased the sensitivity to ABA during seed germination and ABA-mediated stomatal closure. CONCLUSION: Under osmotic stress, CsTLP8 might inhibit seed germination and seedling growth by affecting antioxidant enzymes activities. CsTLP8 acts as a negative regulator in osmotic stress and its effects may be related to ABA.

The complete mitochondrial genome sequence of Fragaria orientalis (Rosaceae).

Fragaria orientalis Lozinsk. is valuable germplasm material for cross breeding in Fragaria. In this study, we assembled the complete mitochondrial genome of F. orientalis using a combination of Illumina data and Nanopore data. The mitochondrial genome was 275,143 bp in length, including 29 protein-coding genes, 20 tRNA genes, and three rRNA genes, with a total GC content 45.23%. Seven protein-coding genes contained introns, and three were trans-spliced. Phylogenetic analysis indicated that F. orientalis is making a sister clade to the Amygdaloideae species. The complete mitochondrial genome of F. orientalis reported in this study will improve our understanding of Fragaria evolution.

Complete chloroplast genome sequencing of ten wild Fragaria species in China provides evidence for phylogenetic evolution of Fragaria.

Complete chloroplast genomes of ten wild Fragaria species native to China were sequenced. Phylogenetic analysis clustered Fragaria species into two clades: The south clade (F. iinumae, F. chinensis, F. pentaphylla, F. nilgerrensis, F. daltoniana, F. corymbosa, F. moupinensis, F. tibetica, F. nipponica, F. gracilis, and F. nubicola and north clade (F. viridis, F. orientalis, F. moschata, F. mandshurica, F. vesca, F. chiloensis, F. virginiana, and F. × ananassa), while F. iinumae is the oldest extant species. Molecular clock analysis suggested present Fragaria species share a common ancestor 3.57 million years ago (Ma), F. moschata and octoploid species evolve 0.89 and 0.97 Ma, respectively, but F. moschata be not directly involved in current octoploid species formation. Drastic global temperature change since the Palaeocene-Eocene, approx. 55 Ma, especially during uplifting of the Qinghai-Tibet plateau and quaternary glaciation may have driven the formation of Fragaria, separation of two groups and polyploidization.

SlTLFP8 reduces water loss to improve water-use efficiency by modulating cell size and stomatal density via endoreduplication.

Improving plant water-use efficiency (WUE) is important to plant survival and crop yield in the context of water limitation. In this study, SlTLFP8 (Tubby-like F-box protein 8) was identified as an osmotic-induced gene in tomato. Transgenic tomato with up-regulated expression of SlTLFP8 showed enhanced water-deficient resistance, whereas knockout mutants generated by CRISPR/Cas9 were more sensitive to water deficit. SlTLFP8 overexpression significantly enhanced WUE by suppressing transpiration under both water-sufficient and water-deficient conditions. Further study showed that overexpressing SlTLFP8 significantly increased leaf epidermal cell size and thereby decreased stomatal density 10-20%, conversely SlTLFP8 knockout resulted in decreased cell size and thereby increased stomatal density 20-50%. SlTLFP8 overexpression and knockout modulated ploidy levels in leaf cells. Changes in expression of cell cycle related genes also indicated that SlTLFP8 affected cell size and stomatal density through endocycle transition. Despite changes in stomata density and transpiration, altering the expression of SlTLFP8 did not change photosynthesis. Additionally, biomass was not altered and there was little difference in fruit yield for transgenic and wild type lines under water-sufficient and water-deficient conditions. Our results demonstrate the effect of SlTLFP8 on endoreduplication and the potential of SlTLFP8 for improvement of WUE. BRIEF SUMMERY: This work found a new mechanism of TLP (Tubby like protein) response to water-deficient stress. SlTLFP8, a member of TLP family, regulates water-deficient resistance by modulating water loss via affecting stomatal density. Expression of SlTLFP8 was induced by osmotic stress. Transgenic tomato lines with SlTLFP8 overexpression or SlTLFP8 knockout showed significantly differences in water-use efficiency (WUE) and water-deficient resistance. The difference of leaf water loss caused by transpiration is the main explanation of the difference in WUE and water-deficient resistance. Additionally, overexpressing SlTLFP8 significantly decreased stomatal density, while SlTLFP8 knockout resulted in increased stomatal density, and SlTLFP8 affected stomatal density through endoreduplication and altered epidermal cell size. Despite changes in stomata density, altering the expression of SlTLFP8 did not result in distinct changes in photosynthesis, biomass and yield of tomato.

Dual Effect of 5-HT1B/1D Receptors on Dopamine Neurons in Ventral Tegmental Area: Implication for the Functional Switch After Chronic Cocaine Exposure.

BACKGROUND: Serotonin (5-HT) 1B/1D receptor (5-HT1B/1DR) agonists undergo an abstinence-induced switch in their effects on cocaine-related behaviors, which may involve changes in modulation of dopamine (DA) neurons in the ventral tegmental area (VTA). However, it is unclear how 5-HT1B/1DRs affect VTA DA neuronal function and whether modulation of these neurons mediates the abstinence-induced switch after chronic cocaine exposure. METHODS: We examined the ability of 5-HT1B/1DRs to modulate D2 autoreceptors (D2ARs) and synaptic transmission in the VTA by slice recording and single unit recording in vivo in naïve mice and in mice with chronic cocaine treatment. RESULTS: We report a bidirectional modulation of VTA DA neuronal firing through the interaction of VTA 5-HT1B/1DRs and D2ARs. In both VTA slices and the VTA of anesthetized mice, the 5-HT1B/1DR agonist CP94253 decreased DA neuronal firing rate and evoked excitatory postsynaptic currents to DA neurons in slice. Paradoxically, CP94253 decreased quinpirole-induced inhibition of DA neurons by reducing D2AR-mediated G protein-gated inwardly rectifying potassium current. This manifested decreased GABAA (gamma-aminobutyric acid A) receptor-mediated evoked inhibitory postsynaptic currents in slice, resulting in disinhibition of DA neurons, in opposition to the 5-HT1B/1DR-induced inhibition. This dual effect was verified in chronic cocaine-treated and mild stress-treated, male mice on days 1 and 20 posttreatment. CONCLUSIONS: This study revealed dual effects of CP94253 on VTA DA neurons that are dependent on D2AR sensitivity, with anti-inhibition under normal D2AR sensitivity and inhibition under low D2AR sensitivity. These dual effects may underlie the ability of CP94253 to both enhance and inhibit cocaine-induced behaviors.

SoHSC70 positively regulates thermotolerance by alleviating cell membrane damage, reducing ROS accumulation, and improving activities of antioxidant enzymes.

High temperature is a major environmental factor affecting plant growth. Heat shock proteins (Hsps) are molecular chaperones that play important roles in improving plant thermotolerance during heat stress. Spinach (Spinacia oleracea) is very sensitive to high temperature; however, the specific function of Hsps in spinach is unclear. In this study, cytosolic heat shock 70 protein (SoHSC70), which was induced by heat stress, was cloned from spinach. Overexpressing SoHSC70 in spinach calli and Arabidopsis enhanced their thermotolerance. In contrast, spinach seedlings with silenced SoHSC70 by virus-induced gene silencing (VIGS) showed more sensitivity to heat stress. Further analysis revealed that overexpressing SoHSC70 altered relative electrical conductivity (REC), malondialdehyde (MDA) content, photosynthetic rate, reactive oxygen species (ROS) accumulation and the activities of antioxidant enzymes, such as superoxide dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX), and catalase (CAT) after the heat treatment. Taken together, our results suggest that overexpressing SoHSC70 positively affects heat tolerance by reducing membrane damage and ROS accumulation and improving activities of antioxidant enzymes.

PvNAC1 increases biomass and enhances salt tolerance by decreasing Na+ accumulation and promoting ROS scavenging in switchgrass (Panicum virgatum L.).

Switchgrass (Panicum virgatum L.) is a bioenergy crop; thus, it is important to improve biomass to effectively produce bioethanol, particularly under adverse stress conditions. NAC transcription factors are involved in the abiotic stress response. PvNAC1 was isolated in the nucleus of switchgrass, with its C-terminal region containing a transcriptional activation domain. PvNAC1 expression was induced by dehydration, salt, H2O2, and abscisic acid treatments. Overexpressing (OE) PvNAC1 improved growth performance, leading to significantly taller and heavier (dry weight) plants. Moreover, cellulose content was significantly higher in OE plants, indicating that PvNAC1 plays an important role regulating growth and bioethanol production. PvNAC1 RNA interference (RNAi) switchgrass plants exhibited reduced dry weight and cellulose content. OE PvNAC1 enhanced tolerance to salt stress, through higher reactive oxygen species scavenging ability and less Na+ and more K+ accumulation in roots and shoots. RNAi plants were more sensitive to salt stress. The quantitative polymerase chain reaction results revealed that some stress responsive genes, three antioxidant enzymatic genes, and an ion homeostasis-related gene were upregulated in OE plants and downregulated in RNAi plants. These results show that PvNAC1 functions as a transcriptional activator in response to salt stress and growth.

Melatonin Alleviates Copper Toxicity via Improving Copper Sequestration and ROS Scavenging in Cucumber.

Melatonin plays an important role in stress tolerance in plants. In this study, exogenous melatonin significantly alleviated the dwarf phenotype and inhibited the decrease of plant fresh weight induced by excess copper (Cu2+). Our results indicated that melatonin alleviated Cu2+ toxicity by improving Cu2+ sequestration, carbon metabolism and ROS (reactive oxygen species) scavenging, rather than by influencing the Cu2+ uptake under excess Cu2+ conditions. Transcriptome analysis showed that melatonin broadly altered gene expression under Cu2+ stress. Melatonin increased the levels of glutathione and phytochelatin to chelate excess Cu2+ and promoted cell wall trapping, thus keeping more Cu2+ in the cell wall and vacuole. Melatonin inhibited ROS production and enhanced antioxidant systems at the transcriptional level and enzyme activities, thus building a line of defense in response to excess Cu2+. The distribution of nutrient elements was recovered by melatonin which was disturbed by Cu2+. In addition, melatonin activated carbon metabolism, especially glycolysis and the pentose phosphate pathway, to generate more ATP, an intermediate for biosynthesis. Taken together, melatonin alleviated Cu2+ toxicity in cucumber via multiple mechanisms. These results will help to resolve the toxic effects of Cu2+ stress on plant growth and development. These results can be used for new strategies to solve problems associated with Cu2+ stress.

CsATAF1 Positively Regulates Drought Stress Tolerance by an ABA-Dependent Pathway and by Promoting ROS Scavenging in Cucumber.

The NAC transcription factors play vital roles in responding to drought stress in plants; however, the molecular mechanisms remain largely unknown in cucumber. Suppression of CsATAF1 via RNA interference (RNAi) weakened drought stress tolerance in cucumber due to a higher water loss rate in leaves, a higher level of hydrogen peroxide (H2O2) and superoxide radicals (O2·-), increased malondialdehyde (MDA) content, lower Fv/Fm ratios and lower antioxidant enzyme activity. The analysis of root length and stomatal apertures showed that CsATAF1-RNAi cucumber plants were less responsive to ABA. In contrast, CsATAF1-overexpression (OE) plants showed increased drought stress tolerance and sensitivity to ABA. Quantitative PCR (qPCR) analysis showed that expression of several stress-responsive genes was significantly up-regulated in CsATAF1-OE transformants and down-regulated in CsATAF1-RNAi transformants. CsABI5, CsCu-ZnSOD and CsDREB2C were verified as direct target genes of CsATAF1. Yeast one-hybrid analysis and electrophoretic mobility shift assay (EMSA) further substantiated that CsATAF1 bound to the promoters of CsABI5, CsCu-ZnSOD and CsDREB2C. Transient expression in tobacco leaves and cucumber protoplasts showed that CsATAF1 directly up-regulated the expression of CsABI5, CsCu-ZnSOD and CsDREB2C. Our results demonstrated that CsATAF1 functioned as a positive regulator in response to drought stress by an ABA-dependent pathway and decreasing reactive oxygen species (ROS) accumulation in cucumber.

Hippocampal synaptic and neural network deficits in young mice carrying the human APOE4 gene.

INTRODUCTION: Apolipoprotein E4 (APOE4) is a major genetic risk factor for late-onset sporadic Alzheimer disease. Emerging evidence demonstrates a hippocampus-associated learning and memory deficit in aged APOE4 human carriers and also in aged mice carrying human APOE4 gene. This suggests that either exogenous APOE4 or endogenous APOE4 alters the cognitive profile and hippocampal structure and function. However, little is known regarding how Apoe4 modulates hippocampal dendritic morphology, synaptic function, and neural network activity in young mice. AIM: In this study, we compared hippocampal dendritic and spine morphology and synaptic function of young (4 months) mice with transgenic expression of the human APOE4 and APOE3 genes. METHODS: Hippocampal dendritic and spine morphology and synaptic function were assessed by neuronal imaging and electrophysiological approaches. RESULTS: Morphology results showed that shortened dendritic length and reduced spine density occurred at hippocampal CA1 neurons in Apoe4 mice compared to Apoe3 mice. Electrophysiological results demonstrated that in the hippocampal CA3-CA1 synapses of young Apoe4 mice, basic synaptic transmission, and paired-pulse facilitation were enhanced but long-term potentiation and carbachol-induced hippocampal theta oscillations were impaired compared to young Apoe3 mice. However, both Apoe genotypes responded similarly to persistent stimulations (4, 10, and 40 Hz for 4 seconds). CONCLUSION: Our results suggest significant alterations in hippocampal dendritic structure and synaptic function in Apoe4 mice, even at an early age.